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Studies of poly-L-lysine-starch nanoparticle preparation and its application as gene carrier

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Science China Chemistry 2005年第2期48卷 162-166页

作者： XIAO Suyao LIU Xuanming TONG Chunyi LIU Jun TANG Dongying ZHAO Lijian Institute of Life Science and Biotechnology College of Chemistry & Chemical Engineering Hunan University Changsha 410082 China Institute of Life Science and Biotechnology College of Chemistry & Chemical Engineering Hunan University Changsha 410082 China Institute of Life Science and Biotechnology College of Chemistry & Chemical Engineering Hunan University Changsha 410082 China Institute of Life Science and Biotechnology College of Chemistry & Chemical Engineering Hunan University Changsha 410082 China Institute of Life Science and Biotechnology College of Chemistry & Chemical Engineering Hunan University Changsha 410082 China Institute of Life Science and Biotechnology College of Chemistry & Chemical Engineering Hunan University Changsha 410082 China

Anion starch nanoparticle (StNP) with a diameter of 50 nm was prepared in wa- ter-in-oil microemulsion, with soluble starch as raw materials and POCl3 as crosslinking agent. PLL-StNP was prepared by linking poly-L-lysine (PLL) on the surface of StNP. At the same time, the size of PLL-StNP and its stability in aqueous solution were checked by AFM. The analysis of plasmid DNA binding, DNase I enzymatic degradation, toxicity and transfection.were done. We discovered that PLL-StNP may be used as non-virus nanoparticle gene carrier. And we devel- oped the method of preparing PLL-StNP gene carrier and used it in cell transfection. As non-virus gene carrier, PLL-StNP has some advantages, such as large load of DNA, high transfection.effi- ciency, low cell toxicity and biodegradability.

关键词： anion starch nanoparticle, poly-L-lysine, non-virus gene carrier, cell transfection.

A novel nonviral nanoparticle gene vector: Poly-L-lysine- silica nanoparticles

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Chinese Science Bulletin 2002年第8期47卷 654-658页

作者： Shiguo Zhu Hongbin Lu Juanjuan Xiang Ke Tang Bicheng Zhang Ming Zhou Chen Tan Guiyuan Li Cent S Univ Xiangya Sch Med Inst Canc Res Changsha 410078 Peoples R China

DNA delivery is a core technology for gene structure and function research as well as clinical settings. The ability to safely and efficiently targeted transfer foreign DNA into cells is a fundamental goal in biotechnology. With the development of nanobiotechnology, nanoparticle gene vectors brought about new hope to reach the goal. In our research, silica nanoparticles (SiNP) were synthesized first in a microemulsion system polyoxyethylene nonylphenyl ether (OP-10)/cyclohexane/ammonium hydroxide, at the same time the effects of SiNP size and its distribution were elucidated by orthogonal analysis; then poly-L-lysine (PLL) was linked on the surface of SiNP by nanoparticle surface energy and electrostatically binding; lastly a novel complex nanomate-rial-poly-L-lysine-silica nanoparticles (PLL-SiNP) wasprepared. The analysis of plasmid DNA binding and DNase I enzymatic degradation discovered that PLL-SiNP could bind DNA, and protect it against enzymatic degradation. Cell transfection.showed that

关键词： nanoparticle gene vector silica nanoparticles poly-L-lysine-silica nanoparticles synthesis DNA delivery cell transfection.

Functional expression of gap junction gene Cx43 and the myogenic differentiation of rhabdomyosarcoma cells

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Science China Chemistry 1995年第3期38卷 305-312页

作者：林仲翔张志谦韩亚玲 C.C.G.Naus K.R.Yu H.Holtzer Department of Cell Biology Beijing Medical School of Oncology Beijing Institute for Cancer ResearchBeijing 100034 China Department of Anatomy University of Western Ontorio London ON N6A5C1Canada Department of Cell and Developmental Biology University of Pennsylvania PA19104 USA

Rhabdomyosarcoma (RD) cells express low levels of the gap junction protein connexin 43 (Cx43), and its mRNA, and display very weak gap junctional intercellular communication (GJIC) as detected by Cx43 immunofluorescence, slot-blot and dye-transfer methods. These cells grow rapidly and show aberrant and incomplete myogenic differentiation. To investigate the role of gap junctions in these cells, the expression of Cx43 with relation to cell growth and myogenic differentiation in RD single-cell subclones-RDL3 and RDL6 is studied. The subclone RDL3 grows slowly and displays better myogenic differentiation. The expression of Cx43, its mRNA and the GJIC in RDL3 is comparable to that in normal myoblasts. Another subclone RDL6 which grows rapidly, but is poorly differentiated, expresses very low levels of Cx43 and its mRNA, and very weak GJIC. By using the calcium phosphate precipitate transfection.technique, a full-length cDNA-encoding Cx43 and a pSV2neo have been introduced into the RDL6 cells. Several

关键词： connexin 43 gap junctional intercellular communication rhabdomyosarcoma myogenic differentiation single-cell clone cell transfection.

Deoxyribozymes inhibit the expression of periodl gene in vitro

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Science China(Life Sciences) 2005年第3期48卷 195-201页

作者： ZHOU Wei WANG Yueqi LIU Yanyou PENG Wenzhen XIAO Jing ZHU Bin WANG Zhengrong Biomedical Engineering Department School of West China Basic and Forensic Science Sichuan University Chengdu610041 China Biomedical Engineering Department School of West China Basic and Forensic Science Sichuan University Chengdu610041 China Biomedical Engineering Department School of West China Basic and Forensic Science Sichuan University Chengdu610041 China Molecular Biology and Biology Chemistry Department School of West China Basic and Forensic Science Sichuan UniversityChengdu610041 China Biomedical Engineering Department School of West China Basic and Forensic Science Sichuan University Chengdu610041 China Biomedical Engineering Department School of West China Basic and Forensic Science Sichuan University Chengdu610041 China Biomedical Engineering Department School of West China Basic and Forensic Science Sichuan University Chengdu610041 China

To investigate the effect of two deoxyribozymes targeting period1 (per1) mRNA in vitro for exploring a novel gene therapy approach about circadian rhythm diseases, the specific deoxyribozymes targeting per1 were designed and synthesized chemically following MFold analysis according to its mRNA secondary structure. per1 RNA fragments were prepared by in vitro transcription of pcDNA3.1(+)-per1164:256. The cleavage reactions containing deoxyribozymes and per1 RNA fragments were performed under certain conditions. With the transfection.tech- nique mediated by LipofectAMINETM, pcDNA3-per1 and DRz164 or DRz256 were introduced into NIH3T3 cells. The effects of deoxyribozymes on per1 were studied by reverse tran- script-polymerase chain reaction (RT-PCR) and flow cytometry (FCM). When deoxyribozymes and RNA transcripts were incubated under the adopted conditions at 37℃ for 2 h, about 63% of per1164:256 RNA transcripts were cleaved by DRz164 and about 50.5% by DRz256. After co- transfecting pcDNA3-per1 with DRz164 or DRz256, the expression of per1 mRNA was de- creased, as indicated by RT-PCR semi-quantity analysis. FCM analysis showed that Per1 protein was inhibited. Both DRz164 and DRz256 targeting per1 have the specific cleavage activity to- ward per1 mRNA in vitro and can highly block the expression of per1 gene in cellular milieu.

关键词： period1 gene, deoxyribozymes, cleavage in vitro, gene transfection.